Method for the early identification and prediction of kidney injury

ABSTRACT

A method for the early identification and prediction of elevated blood creatinine levels resulting from a reduction in kidney function in a subject, comprises contacting a urine sample from the subject with a capture molecule for a biomarker specific for the distal region of the renal tubule and which biomarker is released from said region when there is damage to said region indicative and predictive of elevated blood creatinine levels resulting from a reduction in kidney function. The method can be used to detect Acute Kidney Injury (AKI) caused by many conditions or diseases or through the administration of drugs. The method can indicate and/or predict a reduction in kidney function significantly earlier than the current standard creatinine test. Methods for predicting a need for renal replacement therapy (RRT) are also disclosed.

TECHNICAL FIELD

This invention relates to the early identification and prediction ofkidney damage, including early identification and prediction of elevatedblood creatinine levels resulting from a reduction in kidney function ina subject and, in particular, to biomarkers for the detection thereof.

BACKGROUND ART

There are many causes of a reduction in kidney function and it isessential that corrective action is taken as early as possible byappropriate medical intervention so as to minimise as far as possiblethe deleterious consequences, which include total renal failure and aneed for dialysis and ultimately kidney transplant. For example, anabrupt reduction in kidney function occurs frequently followingcardiothoracic (CT) surgery. Thus, Acute Kidney Injury (AKI) is commonfollowing CT surgery occurring in 7-42% of patients (Mora Mangano, C. etal (1998) Ann Intern Med 128:194-203; and Tuttle, K. R. et al (2003)Amer J. Kid Dis 41:76-83.) Small changes in serum creatinine have beenshown to correlate with increased morbidity and mortality, following CTsurgery (Lassnigg, A. et al (2004) J. Am Soc Nephrol 15; 1597-1605)

Measurement of creatinine is the standard test in the clinic formeasuring kidney function. If kidney function is abnormal, creatininelevels will increase in the blood due to decreased excretion ofcreatinine in the urine. Creatinine levels vary according to a person'sage, size and muscle mass. In acute conditions build up of creatinine inthe blood may take up to 24-72 hours to occur.

Patients who develop severe AKI requiring Renal Replacement Therapy(RRT), after CT surgery have a greatly increased in-hospital mortality(63%) compared to those with non-dialyzed AKI (19%), or stable renalfunction (0.9%) (Mora Mangano, C. et al (1998) supra).

Koyner, J. L. et al (poster presentation at American Society ofNephrology, Renal Week 2007, Oct. 31-Nov. 5, 2007, Moscone Center, SanFrancisco, Calif.) have investigated urinary Cystatin C (CyC) andNeutrophil Gelatinase—Associated Lipocalin (NGAL) in patients with AKIfollowing adult cardiac surgery. Koyner, J L et al found that urinaryCyC excretion increases in the early post-operative period followingadult CT surgery and concluded that urinary CyC may be a useful earlybiomarker for the development of AKI as it appears to correlate with theseverity of AKI and thus the future need of RRT. Similarly, Koyner, J Let al found that urinary NGAL in the early post-operative period appearsto predict the development of AKI and correlate strongly with the futureneed of RRT.

U.S. Publication 2004/0219603 discloses that urinary NGAL measuredwithin two hours of cardiac surgery was predictive of Acute RenalFailure (ARF) as reflected by serum creatinine peak, which occursseveral hours or even days later.

Koyner J. L. et al (2007) (supra) show that for both CyC and NGAL themain increase occurs in the ICU (Intensive Care Unit) phase post CTsurgery.

Eijkenboom, J. J. A. et al (2005) Intensive Care Med 31:664-667 showthat an increase in Glutathione S-Transferase (GST), excretion followingcardiac surgery was not correlated with changes in plasma creatinine andis not associated with clinically relevant renal injury.

Davis, C. L. et al (1999) J Am Soc Nephrol 10: 2396-2402 disclose thaturinary GST excretion increased in most patients after CPB, however,this increase was not associated with the development of clinicallyapparent ARF.

There is a need for a biomarker which predicts elevated blood creatininelevels resulting from an abrupt reduction in kidney function and thusthe development of AKI at the earliest stage post CT surgery, ideally atzero hours in the recovery room and prior to transfer to ICU or earlier,namely intraoperatively, so as to enable corrective action to be takenas soon as possible for those patients who develop AKI with theattendant consequences.

Currently no drug therapy is available for counteracting the effects ofa reduction in kidney function as seen, for example in post CT surgery.Accordingly, the surgeon and other attending medical professionals willendeavour to reduce the effects of renal ischemia or other causativeeffect by managing fluid levels and other physiological parameters.However, as indicated above, frequently, if such measures do not provesuccessful, the patient will require RRT, namely dialysis.

DISCLOSURE OF THE INVENTION

Accordingly, the invention provides in a first aspect a method for theearly identification and prediction of elevated blood creatinine levelsresulting from a reduction in kidney function in a subject, which methodcomprises contacting a urine sample from the subject with a capturemolecule for a biomarker specific for the distal region of the renaltubule and which biomarker is released from said region when there isdamage to said region indicative and predictive of elevated bloodcreatinine levels resulting from a reduction in kidney function.

The method according to the invention, provides a means of detectingdamage to, and predicting the extent of damage to, the kidney within twohours or less of the damage occurring with the attendant advantages forthe patient.

By “capture molecule” herein is meant any molecule or portion thereofwhich binds reversibly or irreversibly to said biomarker, so that saidbiomarker can be detected in the urine sample.

According to one embodiment of the invention, the reduction in kidneyfunction is caused by Acute Kidney Injury (AKI).

According to a further embodiment of the invention, the AKI is caused bya condition or disease selected from age, burns, pre-existing chronickidney disease, reduced effective arterial volume, volume depletion,nephrotic syndrome, congestive heart failure, cirrhosis, sepsis, type Idiabetes, type II diabetes, obesity, inflammation, surgery, solid organtransplant, allogenic bone marrow transplant, mechanical ventilationand/or trauma.

According to a still further embodiment of the invention, the AKI iscaused by administration of a drug to the subject, includingantibiotics.

Preferably, the drug is selected from aminoglycosides, non-steroidalanti-inflammatory drugs and radiocontrast drugs.

It will be appreciated that the AKI may be caused by a toxin.

According to a further embodiment of the invention, the AKI is caused byrenal ischemia in a patient undergoing cardiothoracic (CT) surgery.

Preferably, the biomarker is detectable as early as intraoperatively,allowing for immediate corrective medical intervention.

The method according to the invention, by providing a means of detectingdamage to, and predicting the extent of damage to, the kidney as earlyas intraoperatively represents a very significant advance in themanagement and treatment of patients undergoing CT surgery.

According to one embodiment of the invention, the biomarker isdetectable in the recovery stage post CT surgery, allowing for immediatecorrective medical intervention.

The method according to the invention, by providing a means of detectingdamage to, and predicting the extent of damage to, the kidney in therecovery stage post CT surgery, allows for the appropriate medicalintervention to be taken, dependent on the level of the biomarkerdetected during the recovery stage or earlier, namely intraoperatively.

Thus, the method according to the invention can indicate and/or predicta reduction in kidney function significantly earlier than the currentstandard creatinine test or other current methods hereinabove mentioned.

Preferably, the biomarker is detectable prior to transfer of the patientto the Intensive Care Unit (ICU).

When the abrupt reduction in kidney function is caused by AKI, thereduction in kidney function can be reversed by managing fluid levelsand other physiological parameters.

The abrupt reduction in kidney function may result in a requirement forRenal Replacement Therapy (RRT).

In such a situation, the RRT will generally involve putting the patienton dialysis supplemented, as required, by managing fluid levels andother physiological parameters.

Thus, it will be appreciated that use of the method according to theinvention can result in a significant reduction of the deleterious sideeffects of renal ischemia in a patient undergoing CT surgery.

Preferably, the biomarker is pi glutathione S transferase (πGST), alsoreferred to hereinafter as pi GST.

According to one embodiment, the biomarker is detected by immunoassay.

When the biomarker is πGST, the capture molecule is preferably anantibody to πGST. The antibody may be a monoclonal or a polyclonalantibody which binds to πGST.

For example, the biomarker πGST can be detected using an enzymeimmunoassay, more particularly an Enzyme Linked Immunosorbent Assay(ELISA). In this regard, the πGST can be assayed using a commerciallyavailable kit marketed by Biotrin International Limited, Dublin, Irelandas PI GST EIA, (Catalogue No. BIO 85) which is a 96 well EIA assayformat kit. However, any other conventional assay for detecting πGST canbe used.

It will be appreciated that when the biomarker is πGST, an enzyme, thenthe capture molecule therefor can also be a substrate or co-factortherefor.

Accordingly, according to a further embodiment of the invention, thebiomarker can be detected enzymatically.

According to one embodiment of the invention the biomarker is detectedby a point-of-care assay.

A point-of-care assay will typically be performed on a urine sample ofless than 500 μl, typically 10 μl or less. In a point-of-care assay inaccordance with the invention, the capture medium will be suitably adip-stick or like device having the capture molecule affixed thereto.

The invention also provides πGST for use as a biomarker for the earlyidentification and prediction of elevated blood creatinine levelsresulting from a reduction in kidney function.

According to a further aspect of the invention, there is provided amethod for predicting a need for renal replacement therapy (RRT) in apatient comprising:

determining a concentration of glutathione S transferase (GST) in afirst urine sample from the patient; and

wherein a need for RRT is predicted when the GST concentration isdetermined to be elevated in comparison to a patient without kidneyinjury.

Without being bound by any theoretical explanation of the invention, itis believed that elevated urinary GST concentrations can be used todifferentiate between patients with less severe acute kidney injury notlikely to require RRT and those for whom RRT, for example peritonealdialysis, hemofiltration, renal transplantation and the like, will berequired.

According to this aspect of the invention, the method may furthercomprise contacting a urine sample from the patient with a capturemolecule for a GST isozyme.

According to one embodiment of the invention, the GST is πGST.

The GST can be detected by immunoassay and the capture molecule can bean antibody to πGST.

Alternatively, the GST can be detected enzymatically as in the case ofthe method hereinabove described.

Furthermore, the GST can be detected by a point-of-care assay, ashereinbefore described.

Elevated urinary GST concentrations can persist for days in a patient inneed of RRT.

According to a still further aspect of the invention there is provided amethod for predicting a need for RRT in a patient comprising;

determining a concentration of GST in two urine samples taken at least24 hours apart from the patient; and

wherein a need for RRT is predicted when the GST concentration isdetermined to be elevated in the two urine samples.

The elevated GST concentration in the urine can, for example, be ≧30ng/ml, ≧60 ng/ml, ≧70 ng/ml, ≧80 ng/ml, ≧ or 90 ng/ml or more.

According to this embodiment of the invention, the GST is preferably aπGST isozyme.

The cause of the underlying renal dysfunction in the patient for whomRRT is predicted can, for example, be such that the patient is affectedby an age-related condition, burns, pre-existing chronic kidney disease,reduced effective arterial volume, volume depletion, nephrotic syndrome,congestive heart failure, cirrhosis, sepsis, type I diabetes, type IIdiabetes, obesity, inflammation, surgery, being a solid organ transplantrecipient, being an allogenic bone marrow transplant recipient,mechanical ventilation and/or trauma or has taken or has beenadministered an antibiotic, drug and/or toxin.

The method according to this aspect of the invention can furthercomprises detecting for the presence of risk factors for RRT in thepatient wherein the risk factor is selected from the group consisting ofelevated serum creatinine concentration, type I diabetes, type IIdiabetes, hypertension, dyslipidemia, hyperglycaemia, proteinuria andhypoalbuminemia.

The invention will be described herein with reference to one cause of areduction in kidney function, namely that which frequently occurs in apatient undergoing CT surgery.

Preferably, the biomarker is detected earlier than 2 hours post CTsurgery or earlier than two hours post Cardio-Pulmonary Bypass (CPB).

Further, preferably, the biomarker is detected at zero hours post CTsurgery or CPB.

It will be appreciated that individuals have different urinary biomarkerreference baseline levels. Therefore, post-operative or post-treatmentresults should be considered in relation to the patient's pre-operativeor pre-treatment reference baseline biomarker level, as appropriate.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph of % change in Serum Creatinine (SCr) concentrationfrom baseline versus time as described in Example 1;

FIG. 2. is a graph of absolute change in SCr concentration (mg/dl) frombaseline versus time as described in Example 1;

FIG. 3. is a graph of πGST concentration (ng/ml) versus time asdescribed in Example 1;

FIG. 4. is a graph of πGST concentration (ng/ml) versus time asdescribed in Example 2;

FIG. 5. is a graph of SCr concentration as % of baseline value versustime as described in Example 2; and

FIG. 6. is a graph of absolute change in SCr concentration from baseline(mg/dl) versus time as described in Example 2.

MODES FOR CARRYING OUT THE INVENTION

The invention will be further illustrated by the following Examples

Example 1 Use of πGST as a Biomarker for AKI in Patients Undergoing CTSurgery

A retrospective study of 68 patients who had undergone elective CTsurgery at the University of Chicago Hospital was carried out.

The patients were screened and approached for enrollment. The patientswere excluded if they met any of the following criteria:

Pre-existing End Stage Renal Disease (ESRD) (on RRT) or RenalTransplant.

Age <18 years old.Use of radiocontrast within 24 hours of surgery.Change in thyroid hormone replacement dose in the last 2 weeksChange in thyroid chronic corticosteroids dose in the last 2 weeksUnstable renal function (Δ Serum Creatinine ≧0.2 mg/dl in the last 2months of Oliguria defined as <400 ml/day).

Urine and blood samples were collected and stored.

The urine samples were tested for the presence of πGST using theaforementioned πGST EIA available from Biotrin International Limited(Catalogue Number BIO85).

Serum Creatinine (SCr) was measured using the Jaffe Method in a mannerknown per se on a Beckman Unicel DxC 600 autoanalyser (Beckman Coulter,Fullerton, Calif., USA).

AKI was determined by change in SCr as defined as:

An abrupt (within 48 hours) reduction in kidney function currentlydefined as

1) absolute increase in serum creatinine of more than or equal to 0.3mg/dl (≧26.4 μmol/l); or

2) a percentage increase in serum creatinine of more than or equal to50% (1.5-fold from baseline).

This definition is consistent with the usual definition used, forexample, by Mehta, R. L., et al (2007) Critical Care; 11: R31

The results are shown in Table 1 and FIGS. 1-3

TABLE 1 Future Development of AKI (as defined above) by Day 2 postsurgery AUC* for ROC** Curves & Sensitivity/Specificity at indicatedtime points. AUC Sensitivity Specificity Urinary Pi GST Post Op 0.67963.6% 72.2% % SCr Post Op 0.5 0.0% 100.0% % SCr ICU Admit 0.5 0.0%100.0% % SCr 6 hr post ICU 0.56 12.0% 100.0% % SCr Post Op Day 1 0.7244.0% 100.0% ΔSCr Post Op 0.545 9.1% 100.0% ΔSCr ICU Admit 0.538 7.7%100.0% ΔSCr 6 hr post ICU 0.76 52.0% 100.0% ΔSCr Post Op Day 1 0.8468.0% 100.0% *Area under Curve. *Receiver Operating Characteristic.

FIG. 1. shows the percentage change in SCr from pre-operative baselinevalues for non-AKI patients (--) and AKI patients (-▪-). As shown inFIG. 1, the percentage change in SCr does not increase until after thepatients have been admitted to ICU. However, as AKI is defined as anincrease in SCr of 1.5 fold from baseline, detection of AKI by SCr doesnot occur until Day 2.

FIG. 2. shows the change in absolute value of SCr from pre-operativebaseline values for non-AKI patients (--) and AKI patients (-▪-). Asshown in FIG. 2, a significant increase in SCr concentration does notoccur until 6 hours post ICU in AKI patients. As the definition of AKIis an absolute increase in SCr of more than or equal to 0.3 mg/dl, AKIwould not be diagnosed until after 6 h Post ICU.

FIG. 3. shows urinary πGST levels following CT surgery for non-AKIpatients (--) and AKI patients (-▪-). As shown in FIG. 3, a significantincrease in πGST concentration is observed in Post Op. This indicatedthat patients could be diagnosed with AKI before they are admitted toICU. Although an increase in πGST is observed in non-AKI patients, it issignificantly lower than AKI patient πGST levels, allowing diagnosis ofAKI.

Significantly elevated levels of πGST are detected post-op, namely atzero hours.

Analysis of the data shows that πGST is a good early indicator ofpatients that will develop AKI by day 2 post surgery.

Example 2 Use of πGST as a Biomarker for a Requirement for RRT PatientsUndergoing CT Surgery

A study was carried out on the 68 patients, the subject of Example 1,using the same methodology for the detection of SCr and πGST.

Seven patients out of the 68 patients tested required RRT. The resultsare shown in Table 2.

TABLE 2 Baseline Creatinine Hours in Creatinine at RRT ICU prior (mg/dL)(mg/dL) to RRT Indication 1 5.03 5.4 25.3 Refractory Hyperkalemia (6.0),Oliguria 2 1.49 3.48 51.2 Anuria, Elevated creatinine, Shock 1.36post-op 3 1.3 1.42 21.6 Volume overload, Hypoxia, Oliguria, Hemodynamicinstability *AKI not diagnosed using current SCr measures* 4 1.2 3.7926.8 Lactic Acidosis Oliguria, Shock, Elevated creatinine 5 0.99 1.28 3Lactic Acidosis Anuria, Shock, *AKI not diagnosed using current SCrmeasures* 6 1.19 1.74 5.3 Anuria, Shock (3 pressors), Volume overload.Acidosis 7 1.66 2.8 81 Volume overload, pulmonary edema. Shock

The time point at which patients requiring RRT would be first diagnosedis shown in Table 3.

TABLE 3 SCr increase ≧ 150% SCr increase ≧ 0.3 mg/dL Pi GST > 90 ng/mlAdmit Admit Admit Post- to 6 hr post Post- to 6 hr post Post- to 6 hrpost op ICU ICU Day 1 Day 2 op ICU ICU Day 1 Day 2 op ICU ICU Day 1 Day2 1 Pos Pos 2 Pos Pos Pos 3 Pos 4 Pos Pos Pos 5 Pos * Pos 6 Pos Pos Pos7 Pos * No sample available for testing

Table 4 shows the sensitivity and specificity of πGST to detect RRT assummarised therein.

TABLE 4 Time Cut off point No RRT RRT As determined by Pi GSTconcentration # patients/group  90 ng/ml Post-op No AKI 43 1 AKI 9 3Sensitivity: 83% Specificity: 83% As determined by % SCr increase #patients/group 50% Post-op No AKI 49 5 AKI 0 0 Sensitivity: 0%Specificity: 100% # patients/group 50% Admit to No AKI 60 7 ICU AKI 0 0Sensitivity: 0% Specificity: 100% # patients/group 50% 6 hr post No AKI56 5 ICU admit AKI 2 1 Sensitivity: 17% Specificity: 97% #patients/group 50% Day 1 No AKI 51 4 AKI 8 3 Sensitivity: 43%Specificity: 86% # patients/group 50% Day 2 No AKI 55 4 AKI 4 3Sensitivity: 43% Specificity: 93% As determined by increase of 0.3 mg/dlin SCr # patients/group 0.3 mg/dl Post-op No AKI 47 5 AKI 2 0Sensitivity: 0% Specificity: 96% # patients/group 0.3 mg/dl Admit to NoAKI 58 5 ICU AKI 2 2 Sensitivity: 29% Specificity: 97% # patients/group0.3 mg/dl 6 hr post No AKI 47 3 ICU admit AKI 11 3 Sensitivity: 50%Specificity: 81% # patients/group 0.3 mg/dl Day 1 No AKI 41 3 AKI 18 4Sensitivity: 57% Specificity: 69% # patients/group 0.3 mg/dl Day 2 NoAKI 45 3 AKI 14 4 Sensitivity: 57% Specificity: 76%

The results are also depicted in FIGS. 4-6.

FIG. 4. shows the variation in urinary πGST post CT surgery for non-RRTpatients (--) and RRT patients (-▪-). It will be noted that the πGSTlevel of RRT Patients is significantly higher than non-RRT Patients atthe Post Op time point. FIG. 4 shows a concentration of 135 ng/ml isreached, which is considerably higher than AKI patients shown in FIG. 3(75 ng/ml). This indicates severe AKI and that RRT is required.

FIG. 5. depicts the variation in percentage SCr from baseline post CTsurgery for non-RRT patients (--) and RRT patients (-▪-). FIG. 5 showsthat the percentage change of SCr above baseline is not significantlyelevated above 1.5 fold increase (AKI) until Day 2. This indicates thatthe earliest diagnosis that RRT is required using this technique wouldbe two days following surgery.

FIG. 6. shows the variation in SCr from baseline post CT surgery fornon-RRT patients (--) and RRT patients (--). It will be noted fromFIG. 6 that the absolute change in SCr peaked at Day 2, post surgery. At6 h post ICU a level of 0.3 mg/dl was reached which indicates AKI.Higher concentrations of SCr were measured at Day 1 and Day 2 indicatingsevere AKI and a need for RRT. Using this method, RRT would not beginuntil one day after surgery.

From FIG. 3 and FIG. 4 a relationship is evident between theconcentration of πGST and the damage incurred to the patients' kidneys.A πGST concentration of 300%-500% relative to baseline indicates AKI.However, a πGST concentration greater than 500% indicates severe AKI anda requirement for RRT.

The results show that πGST is a very good early indicator of patientsthat will require RRT undergoing and post CT surgery. * * * * *

The above Examples show that πGST can be used to detect elevated bloodcreatinine, AKI and a requirement for RRT earlier than with currentbiomarkers used to detect an abrupt reduction in kidney function due torenal ischemia intraoperatively or post CT surgery, with the attendantadvantages.

1. A method for the early identification and prediction of elevated blood creatinine levels resulting from a reduction in kidney function in a subject, which method comprises contacting a urine sample from the subject with a capture molecule for a biomarker specific for the distal region of the renal tubule and which biomarker is released from said region when there is damage to said region indicative and predictive of elevated blood creatinine levels resulting from a reduction in kidney function.
 2. A method according to claim 1, wherein the reduction in kidney function is caused by Acute Kidney Injury (AKI).
 3. A method according to claim 2, wherein the AKI is caused by a condition or disease selected from age, burns, pre-existing chronic kidney disease, reduced effective arterial volume, volume depletion, nephrotic syndrome, congestive heart failure, cirrhosis, sepsis, type I diabetes, type II diabetes, obesity, inflammation, surgery, solid organ transplant, allogenic bone marrow transplant, mechanical ventilation and/or trauma.
 4. A method according to claim 2, wherein the AKI is caused by administration of a drug to the subject.
 5. A method according to claim 4, wherein the drug is selected from aminoglycosides, non-steroidal anti-inflammatory drugs and radiocontrast drugs.
 6. A method according to claim 2, wherein the AKI is caused by renal ischemia in a patient undergoing cardiothoracic (CT) surgery.
 7. A method according to claim 6, wherein the biomarker is detectable as early as intraoperatively, allowing for immediate corrective medical intervention.
 8. A method according to claim 6, wherein the biomarker is detectable in the recovery stage post CT surgery, allowing for immediate corrective medical intervention.
 9. A method according to claim 8, wherein the biomarker is detectable prior to transfer of the patient to the Intensive Care Unit (ICU).
 10. A method according to claim 1, wherein the biomarker is pi glutathione S transferase (πGST).
 11. A method according to claim 1, wherein the biomarker is detected by immunoassay.
 12. A method according to claim 11, wherein the capture molecule is an antibody to πGST.
 13. A method according to claim 10, wherein the biomarker is detected enzymatically.
 14. A method according to claim 1, wherein the biomarker is detected by a point-of-care assay.
 15. πGST for use as a biomarker for the early identification and prediction of elevated blood creatinine levels resulting from a reduction in kidney function.
 16. A method for predicting a need for renal replacement therapy (RRT) in a patient comprising: determining a concentration of glutathione S transferase (GST) in a first urine sample from the patient; and wherein a need for RRT is predicted when the GST concentration is determined to be elevated in comparison to a patient without kidney injury.
 17. A method according to claim 16, further comprising contacting a urine sample from the patient with a capture molecule for a GST isozyme.
 18. A method according to claim 16, wherein the GST is πGST.
 19. A method according to claim 16, wherein the GST is detected by immunoassay.
 20. A method according to claim 19, wherein the capture molecule is an antibody to πGST.
 21. A method according to claim 16, wherein the GST is detected enzymatically.
 22. A method according to claim 16, wherein the GST is detected by a point-of-care assay.
 23. A method for predicting a need for RRT in a patient comprising: determining a concentration of GST in two urine samples taken at least 24 hours apart from the patient; and wherein a need for RRT is predicted when the GST concentration is determined to be elevated in the two urine samples.
 24. A method according to claim 16, wherein the elevated GST concentration in urine is ≧30 ng/ml.
 25. A method according to claim 16, wherein the elevated GST concentration in urine is ≧60 ng/ml.
 26. A method according to claim 16, wherein the elevated GST concentration in urine is ≧70 ng/ml.
 27. A method according to claim 16, wherein the elevated GST concentration in urine is ≧80 ng/ml.
 28. A method according to claim 16, wherein the elevated GST concentration in urine is ≧90 ng/ml or more.
 29. A method according to claim 16, wherein the GST is a πGST isozyme.
 30. A method according to claim 16, wherein the patient is affected by an age-related condition, burns, pre-existing chronic kidney disease, reduced effective arterial volume, volume depletion, nephrotic syndrome, congestive heart failure, cirrhosis, sepsis, type I diabetes, type II diabetes, obesity, inflammation, surgery, being a solid organ transplant recipient, being an allogenic bone marrow transplant recipient, mechanical ventilation and/or trauma or has taken or has been administered an antibiotic, drug and/or toxin.
 31. A method according to claim 16, further comprising detecting for the presence of risk factors for RRT in the patient wherein the risk factor is selected from the group consisting of elevated serum creatinine concentration, type I diabetes, type II diabetes, hypertension, dyslipidemia, hyperglycaemia, proteinuria and hypoalbuminemia. 